The yeast mitochondrial citrate transport protein: Determination of secondary structure and solvent accessibility of transmembrane domain IV using site-directed spin labeling

被引:18
|
作者
Kaplan, RS
Mayor, JA
Kotaria, R
Walters, DE
Mchaourab, HS
机构
[1] Finch Univ Hlth Sci Chicago Med Sch, Dept Biochem & Mol Biol, N Chicago, IL 60064 USA
[2] Med Coll Wisconsin, Biophys Res Inst, Natl Biomed Esr Ctr, Milwaukee, WI 53226 USA
关键词
D O I
10.1021/bi000433e
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
To explore the spatial organization and functional dynamics of the citrate transport protein (CTP), a nitroxide scan was carried out along 22 consecutive residues within the fourth transmembrane domain (TMDIV). This domain has been implicated as being of unique importance to the CTP mechanism due to (i) the presence of two intramembranous positive charges that are essential for CTP function and (ii) the existence of a transmembrane aqueous surface within this domain which likely corresponds to a portion of the citrate translocation pathway. The sequence-specific variation in the mobilities of the introduced nitroxides and their accessibilities to molecular O-2 reveal an alpha-helical conformation along the sequence. The accessibilities to NiEDDA are out of phase with accessibilites to O-2, indicating that one face of the helix is solvated by the lipid bilayer while the other is solvated by an aqueous environment. A gradient of NiEDDA accessibility is observed along the helix surface facing the aqueous phase, and the EPR spectral line shapes at these sites indicate considerable motional restriction. In the context of the model where TMDIV lines the translocation pathway, these data suggest a barrier to passive diffusion through the pathway. This paper reports the first use of site-directed spin labeling to study mitochondrial transporter structure.
引用
收藏
页码:9157 / 9163
页数:7
相关论文
共 50 条
  • [21] Membrane protein structure and dynamics studied by site-directed spin-labeling ESR
    Bordignon, Enrica
    Steinhoff, Heinz-Jurgen
    ESR SPECTROSCOPY IN MEMBRANE BIOPHYSICS, 2007, 27 : 129 - +
  • [22] The yeast mitochondrial citrate transport protein - Characterization of transmembrane domain III residue involvement in substrate translocation
    Ma, CL
    Kotaria, R
    Mayor, JA
    Remani, S
    Walters, DE
    Kaplan, RS
    JOURNAL OF BIOLOGICAL CHEMISTRY, 2005, 280 (03) : 2331 - 2340
  • [23] Structure of the substrate binding pocket of the multidrug transporter EmrE: Site-directed spin labeling of transmembrane segment 1
    Koteiche, HA
    Reeves, MD
    Mchaourab, HS
    BIOCHEMISTRY, 2003, 42 (20) : 6099 - 6105
  • [24] Site-directed spin labeling demonstrates that transmembrane domain XII in the lactose permease of Escherichia coli is an alpha-helix
    Voss, J
    He, MM
    Hubbell, WL
    Kaback, HR
    BIOCHEMISTRY, 1996, 35 (39) : 12915 - 12918
  • [25] Solution structure of the cytoplasmic domain of erythrocyte membrane band 3 determined by site-directed spin labeling
    Zhou, Z
    DeSensi, SC
    Stein, RA
    Brandon, S
    Dixit, M
    McArdle, EJ
    Warren, EM
    Kroh, HK
    Song, LK
    Cobb, CE
    Hustedt, EJ
    Beth, AH
    BIOCHEMISTRY, 2005, 44 (46) : 15115 - 15128
  • [26] Estimation of binding parameters for the protein–protein interaction using a site-directed spin labeling and EPR spectroscopy
    Marcin Sarewicz
    Sebastian Szytuła
    Małgorzata Dutka
    Artur Osyczka
    Wojciech Froncisz
    European Biophysics Journal, 2008, 37 : 483 - 493
  • [27] Probing the accessibility of engineered cysteine residues within transmembrane domain III of the mitochondrial citrate transport protein to a methanethiosulfonate derivative
    Kaplan, RS
    Ma, CL
    Mayor, JA
    Walters, DE
    Kotaria, R
    BIOPHYSICAL JOURNAL, 2002, 82 (01) : 568A - 568A
  • [28] Structure of the KcsA potassium channel from Streptomyces lividans:: A site-directed spin labeling study of the second transmembrane segment
    Gross, A
    Columbus, L
    Hideg, K
    Altenbach, C
    Hubbell, WL
    BIOCHEMISTRY, 1999, 38 (32) : 10324 - 10335
  • [29] INVESTIGATION OF STRUCTURE AND DYNAMICS IN MEMBRANE-PROTEINS USING SITE-DIRECTED SPIN-LABELING
    HUBBELL, WL
    ALTENBACH, C
    CURRENT OPINION IN STRUCTURAL BIOLOGY, 1994, 4 (04) : 566 - 573
  • [30] Applications of Genetically Encoded Unnatural Amino Acids for Protein Structure Mapping by Site-Directed Spin Labeling
    Fleissner, Mark R.
    Bridges, Michael D.
    Kalai, Tamas
    Axup, Jun Y.
    Young, Douglas D.
    Hideg, Kalman
    Schultz, Peter G.
    Hubbell, Wayne L.
    BIOPHYSICAL JOURNAL, 2011, 100 (03) : 51 - 51